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Study on Vitis Somatic Embryo Induction and Chinese Wild Vitis Stilbene Syntase Gene VpSTSgDAN2 Transformation

Author: XiaoYu
Tutor: WangYueJin
School: Northwest University of Science and Technology
Course: Of Horticultural Plant Germplasm Resources
Keywords: Grapes Somatic embryogenesis Agrobacterium-mediated Stilbene synthase Promoter
CLC: S663.1
Type: Master's thesis
Year: 2011
Downloads: 49
Quote: 0
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Stilbene synthase (stilbene synthase, STS) gene and its specific promoter-based research group pre-associated genes in Chinese wild the Huadong grapes Shirakawa -35 -1 cloned disease resistance. In this study placed to add the stilbene synthase gene enhancer promoter, to build four plant expression vector, aimed at using genetic engineering techniques, it is transferred to a sense of the disease in European grape genome, acquired resistance improve the transgenic plants to provide theoretical and technical basis. 'Jiali stuffed', 'Thompson Seedless', 'Flame Seedless', 'long spike Seedless', 'Cabernet Sauvignon' floral organ explant somatic embryogenesis and seedling research based on Agrobacterium-mediated transformation, will carry the plant expression vector of wild grape stilbene synthase gene-specific promoter into the tomatoes and grapes, the main results obtained are as follows: 1. 5 kinds of European grapes varieties 'Carignan', 'Thompson Seedless', 'Flame Seedless', 'the long spike Seedless', 'Cabernet Sauvignon' stamen, pistil and small buds as explants cultured in four different based on the induction of callus, which medium NN69 2.0 mg · L -1 6-BA 1.0 mg · L -1 2,4-D, 60 g · L -1 sucrose 3 g · L -1 phatagel and medium MS 0.55 mg · L -1 2,4-D 0.5 mg · L -1 NOA 1.24 mg · L -1 4-CPPU 30 g · L -1 sucrose 3 g · L -1 phatagel floral organ explants embryogenic callus induction favorable, the induction rate up to 6.2%. Studied five European grape varieties, in addition to the 'Cabernet Sauvignon', and the remaining four species by the floral organs induced callus available in medium X6 60 g · L -1 sucrose, 7 g L -1 the TC-Agar 0.5 g · L -1 AC further induced somatic embryos. And X6 medium relay generation to save for the developmental stages of somatic embryos before the heart-type embryos to produce secondary embryos. Somatic embryos in liquid MS medium 110rpm 25 ℃ the dark culture 1w transfer to a Tony medium WPM 0.15 mg · L -1 IBA 30 g · L -1 < placed under strong light training / sup> sucrose g · L -1 agar 0.5 g · L -1 AC, the first low-light for 3 d, 1 w The left and right to seedling. 4 by Agrobacterium mediated Law, Wildvine stilbene synthase gene and its specific promoter containing vectors were transfected into tomato Micro-Tom, 51 transgenic seedlings were obtained by the preliminary detection of PCR and PCR-Southern. By high performance liquid chromatography of tomato transgenic plants and wild-type plants inoculated with Botrytis cinerea 24 h after resveratrol content determination, the results indicated the presence of transgenic plants with the highest content of resveratrol enhanced element Enhancer, is not to add any enhancement element vector gD2 into tomato 4.4 times the resveratrol content. Wild-type plants were not detected in resveratrol. The inoculated tomato gray mold disease index was measured, and the results show that the transgenic tomato have some resistance, which contain enhanced disease strongest element Enhancer enhanced vector transformed tomato.

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